本工作由瑞士国家科学基金会（ - 智能材料NRP 62）的支持。也承认是B HANSELMANN，K.凯尔，U.茨和B Leuthold的支持。

1。等离子体引发聚合<ol><li>准备的单体溶液。 <ol><li>在200毫升水中，并用己烷（100毫升）的分液漏斗中洗涤3次，溶解HEMA（0.718摩尔100ml）中。饱和水相用氯化钠和提取的HEMA，用乙醚（50毫升）中。干燥有机相用MgSO 4和除去溶剂在真空中 （100毫巴，40℃）。蒸馏减压HEMA（15毫巴; 99​​℃）。 </li><li>准备在第1.1.1节中产生的无抑制剂的HEMA的0.62 2M甲醇溶液。倾30毫升溶液的成1颈烧瓶中，并通过1小时的溶液中鼓入氩气排除氧。 </li></ol></li><li>表面引发聚合。 <ol><li>位置2的聚碳酸酯膜彼此相邻到等离子体腔室（ 图2）。将膜朝向气相的光泽面。 </li><li>等离子腔连接到啊IGH真空（20毫巴），持续5分钟。关闭阀至真空，并打开另一个阀，其连接到氩气和氧气的气体和2小时15 sccm的氩气和2.5 sccm的氧气吹扫腔室与该混合物。 </li><li>启动等离子体，并减少功率到所需的功率（用于聚碳酸酯膜：12瓦）和治疗的膜为4分钟，等离子体。通过打开相应的阀连接的单体溶液与腔室。关闭血浆和疏散室。 </li><li>通过打开相应的阀连接的单体溶液与所述腔室和将溶液倾入腔室。确保该膜覆盖与单体溶液。打开连接在氩气阀门和12小时存储该反应混合物在20°C（空调房间）。 </li><li>除去单体溶液。用甲醇在超声浴中5分钟，洗膜。用水重复洗涤过程。 </li><li>干燥该膜在真空下在分子筛上2小时。 </li></ol></li></ol> 2。涂层膜的修改后的<ol><li>准备spirobenzopyran 1的溶液（ 图1）（100毫克，0.27毫摩尔），在叔 butylmetylether和二甲基氨基吡啶（DMAP）（33毫克，0.27毫摩尔），N，N-二环己基碳（DCC）（0.27毫摩尔55毫克） （TBME）（12毫升）中。 </li><li>放置一保护搅拌棒和一个保护网格成一个圆底烧瓶中。干燥瓶和洪水量瓶中，用氩气。 </li><li>将溶液倒入烧瓶，随后涂覆的隔膜。 </li><li>轻轻搅拌在室温下搅拌12小时。 </li><li>除去溶液，并用叔 butylmetylether洗膜在超声浴中5分钟。用乙醇和水重复洗涤过程。 </li><li>干燥该膜在真空中在分子筛上2小时。 </li></ol> 3。表面张力测试<ol><l我&gt;修复膜与金属O型圈标准的磁带。定位超纯水（3.3微升）的在膜表面上，这是不与O形圈接触的那一部分的下降。测量接触角（CA）在膜的5个不同的斑点。 </li><li>用于测试样品的长期稳定性，测量接触角在三个不同的膜的斑点0后，1，2，3，7，14，和21天。 </li></ol> 4。光致变色膜的透气性测试<ol><li>装满水的Franz扩散池（12毫升）的受体室。 </li><li>修复膜在Franz扩散池。确保该膜是在与受体腔室中的水接触。用含水咖啡因溶液（;3.0毫升20毫摩尔）填充供料腔室（在膜的顶部的腔室）。从供体室用白色光（ 图3）的顶部照射该膜。从受体细胞收集样品（200微升）; FO用200nm的细孔孔径r轨道边的聚碳酸酯膜，收集样品每10分钟。 </li><li>如重复步骤4.2所描述的实验。但照射整个渗透性试验过程中，用紫外灯（366纳米，80瓦/米2）的膜。 </li><li>测定采集的样本的咖啡因浓度。 <ol><li>绘制校准曲线与使用的UV / Vis光谱仪15个不同的咖啡因浓度（0.05毫克/毫升和1.5mM / L）。校准在293 nm处。 </li><li>确定每个使用校正曲线所收集的样品的浓度。 </li><li>画出所确定的浓度比 。所收集的样本的时间。做线性拟合，通过点，并确定ΔC从斜坡。 </li></ol></li></ol>

<ol>
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作者什么都没有透露。

等离子体工艺产生紫色气体，该气体是由离子化的氩气。橙色则表示不希望的氮气从泄漏的存在。等离子体处理不只是在表面上形成自由基而且蚀刻膜7,12。太多的蚀刻可以显著改变孔径，这将影响到膜的渗透性。所提出的设置的受控反应条件允许提高等离子体引发的接枝方法的再现性。然而，在等离子体腔室中的膜的确切位置仍然可以在表面上形成自由基的密度影响由于等离子体的不均?...

材料的等离子体改性已成为许多工业领域一个重要的过程。表面和表面官能化，而 ​​不改变该材料的整体性质的清洗已在等离子体处理在表面科学1-8必不可少的过程。 聚合物的等离子处理结果在均裂键的断裂。这导致了聚合物材料的一个边和自由基丰富的表面的形成。通过采用等离子体含有氧分子，表面变得富氧从而更亲水9-11。然而，表面的亲水性并不稳定随着时间的推移12。为了提高长期稳定性，等离子体处理的表面可进行化学后或在等离子体处理13-15修改。通常通过添加反应性单体物种进入气相中的等离子体处理进行该处理;这些单体聚合，然后从聚合物表面产生自由基。如果与一个非易失性单体进行化学处理，该聚合物接枝有等离子体改性后发生。为了执行控制接枝基团在表面上形成后，等离子体的设置被描述，其允许从所述表面在溶液中在控制条件下12,16等离子体引发的表面诱导聚合。 这次报告的重点是轨道边聚合物膜12,17的修改。通过修改这些膜的表面张力，透气率可变化12。这种清洁和快速的过程可以创建非常薄的层（&lt;5 nm）的，涵盖整个膜表面不改变聚合物膜的大部分财产。由于在等离子体处理磨边，轨迹一柄膜的孔径增大略12。磨边率dependi纳克的聚合物上，并具有线性特性。 当使用具有反应性官能团的单体，接枝聚合物可进一步官能化。这表现在一个PHEMA接枝膜用羧酸功能化的螺吡喃的修改后的。这导致光致变色表面，由于螺吡喃已知变换成部花青类时用UV光照射。螺吡喃形式可以通过照射部花青形式与可见光（ 图1）18,19重新建立。由于花青形式比螺吡喃状态更极性的，该涂层的表面张力可与光20被触发。在表面张​​力的变化会影响对水溶液的膜的渗透性阻力。的设立如何执行这些光响应膜的渗透性测试将显示在阻透性的显著变化（减少我97％不阻透性）是证明。这样的膜可以被集成在一个药物递送的设置或在智能传感系统。 <img alt="图1" fo:content-width="5in" fo:src="/files/ftp_upload/51680/51680fig1highres.jpg" src="/files/ftp_upload/51680/51680fig1.jpg" /> 图1。光异构化spirobenzopyran化合物1。

<table border="0" cellpadding="0" cellspacing="0" style="width:943px;" width="943">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Name of Reagent/ Equipment</td>
			<td style="width:284px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">2-Hydroxyethyl methacrylate, 97%</td>
			<td style="width:284px;">Sigma-Aldrich</td>
			<td style="width:119px;">128635</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Hexane 99%</td>
			<td style="width:284px;">Biosolve</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Magnesium sulfate (MgSO4, anhydrous)</td>
			<td style="width:284px;">Sigma-Aldrich</td>
			<td>M7506</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Methanol, 99%&#160;</td>
			<td style="width:284px;">Sigma-Aldrich</td>
			<td style="width:119px;">14262</td>
			<td>dried over molecular sieves</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">N,N-Dicylcohexylcarbodiimide, 99%</td>
			<td style="width:284px;">Sigma-Aldrich</td>
			<td style="width:119px;">D8002</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Dimethyl aminopyridine, 99%</td>
			<td style="width:284px;">Sigma-Aldrich</td>
			<td style="width:119px;">107700</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Tert-butylmethylether, 98%</td>
			<td style="width:284px;">Fluka</td>
			<td style="width:119px;">306975</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Polycarbonate membrane</td>
			<td style="width:284px;">Whatman</td>
			<td style="width:119px;"></td>
			<td>Nanopore Track Etched (TE) (1.0 &#956;m, 0.2 &#956;m, 0.1 &#956;m, 50 nm, 30 nm and 15 nm pore diameter; 47 mm or 25 mm membrane diameter)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Caffeine (reagent plus)</td>
			<td style="width:284px;">Sigma-Aldrich</td>
			<td style="width:119px;">C0750&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Franz diffusion cell (12 ml)</td>
			<td style="width:284px;">SES-Analysesysteme</td>
			<td style="width:119px;">6C010015</td>
			<td>15mm unjacheted Franz Cell, 12 ml Receptor volume, Flat ground, clear glass, stirbat and clamp</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">UV-Lamp</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td>UV irradiation (366 nm, 15 W/m2)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">White light lamp</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td>White light irradiation (500 W bulb)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">UV/Vis spectrophotometer</td>
			<td style="width:284px;">Varian 50Bio/50MPR</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:373px;">Polyester membranes</td>
			<td style="width:284px;">Sterlitech</td>
			<td style="width:119px;">PET0225100</td>
			<td>Polyester Membrane Filters, 0.2 &#956;m pore diameter, 25 mmm diameter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Polyvinylidene fluoride membranes</td>
			<td style="width:284px;">Millipore</td>
			<td style="width:119px;"></td>
			<td>PVDF Membranes Durapore (0.22 &#956;m pore diameter; 47 mm membrane diameter)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Argon (99.9995%)</td>
			<td style="width:284px;">Alphagaz</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Dressler Cesar RF Power Generator</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td>Plasma chamber setup</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">MKS Multi Gas Controller 647C</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td>Plasma chamber setup</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">MKS Mass-Flow controllers</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td>Plasma chamber setup</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Vacuubrand RE 2.5 rotary vane vacuum pump</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td>Plasma chamber setup</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Contact angle measurement device Kr&#252;ss G10</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Balances Mettler Toledo AB204-S and Mettler ME30</td>
			<td style="width:284px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

preparation of light-responsive membranes by a combined surface grafting and postmodification process

为了修改商业可用的轨道边缘的聚合物膜的表面张力，表面引发聚合反应的过程被呈现。从膜表面的聚合反应是由等离子体处理的膜中，接着在膜表面具有2 - 羟乙酯（HEMA）的甲醇溶液反应诱导。特别注意的是考虑到工艺参数对等离子体处理的表面上的聚合反应之前。例如，等离子体处理对不同类型的膜（ 例如聚酯，聚碳酸酯，聚偏二氟乙烯）的影响进行了研究。另外，表面接枝膜的依赖于时间的稳定性示出由接触角测量。当接枝聚（2 - 羟乙基甲基丙烯酸酯）（PHEMA），以这种方式，表面可进一步通过与所需物质的羧酸功能的聚合物的醇部分酯化改性。这些反应，因此可用于对膜表面的官能化。例如，在膜的表面张力可以改变或可以被插入期望的功能所呈现的光响应性。这表现在与这导致了光响应性膜的羧酸官能spirobenzopyran单元反应PHEMA。溶剂的选择起着在修改后的工序的主要作用，并更详细地在本文中进行了讨论。这样的官能化膜的渗透性测量使用Franz池与外部光源进行的。通过改变光的波长从可见光到紫外光范围内，水的咖啡因溶液的渗透性的变化被观察到。

刻蚀速率可以跟随不同的时间段后称量该膜。如可以从图4中可以看出，蚀刻速率如下聚酯，聚偏二氟乙烯，和聚碳酸酯膜的线性刻蚀速率，这可从蚀刻时间与重量损失的线性相关的斜率来确定。 如图4所示 ，聚碳酸酯膜显示所有三个聚合物膜的最低蚀刻速率。该蚀刻的结果之一是在孔径的变化。孔的后等离子体处理增加了约20％12,17的直径。随后的接枝聚?...

Watch this Scientific Journal Video about Preparation of Light-responsive Membranes by a Combined Surface Grafting and Postmodification Process at JoVE.com

Preparation of Light-responsive Membranes by a Combined Surface Grafting and Postmodification Process

一种等离子体诱导聚合反应过程被描述为在聚合物膜的表面引发聚合反应。与光致变色物质的接枝聚合物的进一步修改后的呈现与开展的光响应膜渗透性测量的协议。

通过结合表面接枝和修改后的过程光敏感膜的制备

In order to modify the surface tension of commercial available track-edged polymer membranes, a procedure of surface-initiated polymerization is ...

preparation-light-responsive-membranes-combined-surface-grafting

Research

JoVE Journal

Bioengineering

11.7K 次观看.  Empa, Swiss Federal Laboratories for Materials Science and Technology. 一种等离子体诱导聚合反应过程被描述为在聚合物膜的表面引发聚合反应。与光致变色物质的接枝聚合物的进一步修改后的呈现与开展的光响应膜渗透性测量的协议。 

视频: Preparation of Light-responsive Membranes by a Combined Surface Grafting and Postmodification Process

Measurement of Aggregate Cohesion by Tissue Surface Tensiometry

Rigorous measurement of intercellular binding energy can only be made using methods grounded in thermodynamic principles in systems at equilibrium. We have developed tissue surface tensiometry (TST) specifically to measure the surface free energy of interaction between cells. The biophysical concepts underlying TST have been previously described in detail1,2. The method is based on the observation that mutually cohesive cells, if maintained in shaking culture, will spontaneously assemble into clusters. Over time, these clusters will round up to form spheres. This rounding-up behavior mimics the behavior characteristic of liquid systems. Intercellular binding energy is measured by compressing spherical aggregates between parallel plates in a custom-designed tissue surface tensiometer. The same mathematical equation used to measure the surface tension of a liquid droplet is used to measure surface tension of 3D tissue-like spherical aggregates. The cellular equivalent of liquid surface tension is intercellular binding energy, or more generally, tissue cohesivity. Previous studies from our laboratory have shown that tissue surface tension (1) predicts how two groups of embryonic cells will interact with one another1-5, (2) can strongly influence the ability of tissues to interact with biomaterials6, (3) can be altered not only through direct manipulation of cadherin-based intercellular cohesion7, but also by manipulation of key ECM molecules such as FN8-11 and 4) correlates with invasive potential of lung cancer12, fibrosarcoma13, brain tumor14 and prostate tumor cell lines15. In this article we will describe the apparatus, detail the steps required to generate spheroids, to load the spheroids into the tensiometer chamber, to initiate aggregate compression, and to analyze and validate the tissue surface tension measurements generated.

We describe a method of measuring binding energy, expressible as tissue surface tension, between cells within 3D tissue-like aggregates. Differences in tissue surface tension have been demonstrated to correlate with invasiveness of lung, muscle, and brain tumors, and are fundamental determinants of establishing spatial relationships between different cell types.

总结凝聚力测量组织表面Tensiometry

Rigorous measurement of intercellular binding energy can only be made using methods grounded in thermodynamic principles in systems at equilibrium.  We ...

科研

生物工程

Self-reporting Scaffolds for 3-Dimensional Cell Culture

Culturing cells in 3D on appropriate scaffolds is thought to better mimic the in vivo microenvironment and increase cell-cell interactions. The resulting 3D cellular construct can often be more relevant to studying the molecular events and cell-cell interactions than similar experiments studied in 2D. To create effective 3D cultures with high cell viability throughout the scaffold the culture conditions such as oxygen and pH need to be carefully controlled as gradients in analyte concentration can exist throughout the 3D construct. Here we describe the methods of preparing biocompatible pH responsive sol-gel nanosensors and their incorporation into poly(lactic-co-glycolic acid) (PLGA) electrospun scaffolds along with their subsequent preparation for the culture of mammalian cells. The pH responsive scaffolds can be used as tools to determine microenvironmental pH within a 3D cellular construct. Furthermore, we detail the delivery of pH responsive nanosensors to the intracellular environment of mammalian cells whose growth was supported by electrospun PLGA scaffolds. The cytoplasmic location of the pH responsive nanosensors can be utilized to monitor intracellular pH (pHi) during ongoing experimentation.

Biocompatible pH responsive sol-gel nanosensors can be incorporated into poly(lactic-co-glycolic acid) (PLGA) electrospun scaffolds. The produced self-reporting scaffolds can be used for in&#160;situ monitoring of microenvironmental conditions whilst culturing cells upon the scaffold. This is beneficial as the 3D cellular construct can be monitored in real-time without disturbing the experiment.

自我报告的支架3维细胞培养

Culturing cells in 3D on appropriate scaffolds is thought to better mimic the in vivo microenvironment and increase cell-cell interactions. The resulting ...

Generation and Grafting of Tissue-engineered Vessels in a Mouse Model

The construction of vascular conduits is a fundamental strategy for surgical repair of damaged and injured vessels resulting from cardiovascular diseases. The current protocol presents an efficient and reproducible strategy in which functional tissue engineered vessel grafts can be generated using partially induced pluripotent stem cell (PiPSC) from human fibroblasts. We designed a decellularized vessel scaffold bioreactor, which closely mimics the matrix protein structure and blood flow that exists within a native vessel, for seeding of PiPSC-endothelial cells or smooth muscle cells prior to grafting into mice. This approach was demonstrated to be advantageous because immune-deficient mice engrafted with the PiPSC-derived grafts presented with markedly increased survival rate 3 weeks after surgery. This protocol represents a valuable tool for regenerative medicine, tissue engineering and potentially patient-specific cell-therapy in the near future.

Here, we present a protocol to generate tissue engineered vessel grafts that are functional for grafting into mice by double seeding partially induced pluripotent stem cell (PiPSC) - derived smooth muscle cells and PiPSC - derived endothelial cells on a decellularized vessel scaffold bioreactor.

组织工程血管在小鼠模型的生成和嫁接

The construction of vascular conduits is a fundamental strategy for surgical repair of damaged and injured vessels resulting from cardiovascular diseases. ...

Adapting the Electrospinning Process to Provide Three Unique Environments for a Tri-layered In Vitro Model of the Airway Wall

Electrospinning is a highly adaptable method producing porous 3D fibrous scaffolds that can be exploited in in vitro cell culture. Alterations to intrinsic parameters within the process allow a high degree of control over scaffold characteristics including fiber diameter, alignment and porosity. By developing scaffolds with similar dimensions and topographies to organ- or tissue-specific extracellular matrices (ECM), micro-environments representative to those that cells are exposed to in situ can be created. 
The airway bronchiole wall, comprised of three main micro-environments, was selected as a model tissue. Using decellularized airway ECM as a guide, we electrospun the non-degradable polymer, polyethylene terephthalate (PET), by three different protocols to produce three individual electrospun scaffolds optimized for epithelial, fibroblast or smooth muscle cell-culture. Using a commercially available bioreactor system, we stably co-cultured the three cell-types to provide an in vitro model of the airway wall over an extended time period.
This model highlights the potential for such methods being employed in in vitro diagnostic studies investigating important inter-cellular cross-talk mechanisms or assessing novel pharmaceutical targets, by providing a relevant platform to allow the culture of fully differentiated adult cells within 3D, tissue-specific environments.

Adapting the Electrospinning Process to Provide Three Unique Environments for a Tri-layered In Vitro Model of the Airway Wall

Advancements in biomaterial technologies enable the development of three-dimensional multi-cell-type constructs. We have developed electrospinning protocols to produce three individual scaffolds to culture the main structural cells of the airway to provide a 3D in vitro model of the airway bronchiole wall.

适应静电过程中提供三个独特的环境为一个三层次<em&gt;在体外</em&gt;的气道壁模型

Electrospinning is a highly adaptable method producing porous 3D fibrous scaffolds that can be exploited in in vitro cell culture. Alterations to ...

Qualitative Characterization of the Aqueous Fraction from Hydrothermal Liquefaction of Algae Using 2D Gas Chromatography with Time-of-flight Mass Spectrometry

Two-dimensional gas chromatography coupled with time-of-flight mass spectrometry is a powerful tool for identifying and quantifying chemical components in complex mixtures. It is often used to analyze gasoline, jet fuel, diesel, bio-diesel and the organic fraction of bio-crude/bio-oil. In most of those analyses, the first dimension of separation is non-polar, followed by a polar separation. The aqueous fractions of bio-crude and other aqueous samples from biofuels production have been examined with similar column combinations. However, sample preparation techniques such as derivatization, solvent extraction, and solid-phase extraction were necessary prior to analysis. In this study, aqueous fractions obtained from the hydrothermal liquefaction of algae were characterized by two-dimensional gas chromatography coupled with time-of-flight mass spectrometry without prior sample preparation techniques using a polar separation in the first dimension followed by a non-polar separation in the second. Two-dimensional plots from this analysis were compared with those obtained from the more traditional column configuration. Results from qualitative characterization of the aqueous fractions of algal bio-crude are discussed in detail. The advantages of using a polar separation followed by a non-polar separation for characterization of organics in aqueous samples by two-dimensional gas chromatography coupled with time-of-flight mass spectrometry are highlighted.

A two-dimensional gas chromatography-time-of-flight mass spectrometry method is described for characterization of the aqueous fraction of bio-crude produced from hydrothermal liquefaction of algae. This protocol can also be employed to analyze the aqueous fraction of liquid products from fast pyrolysis, catalytic fast pyrolysis, catalytic deoxygenation and hydro-treating.

从藻类水热液化使用二维气相色谱时间飞行质谱法水提取部分的定性鉴定

Two-dimensional gas chromatography coupled with time-of-flight mass spectrometry is a powerful tool for identifying and quantifying chemical components in ...

Automated Robotic Dispensing Technique for Surface Guidance and Bioprinting of Cells

This manuscript describes the introduction of cell guidance features followed by the direct delivery of cells to these features in a hydrogel bioink using an automated robotic dispensing system. The particular bioink was selected as it allows cells to sediment towards and sense the features. The dispensing system bioprints viable cells in hydrogel bioinks using a backpressure assisted print head. However, by replacing the print head with a sharpened stylus or scalpel, the dispensing system can also be employed to create topographical cues through surface etching. The stylus movement can be programmed in steps of 10 &#181;m in the X, Y and Z directions. The patterned grooves were able to orientate mesenchymal stem cells, influencing them to adopt an elongated morphology in alignment with the grooves&#39; direction. The patterning could be designed using plotting software in straight lines, concentric circles, and sinusoidal waves. In a subsequent procedure, fibroblasts and mesenchymal stem cells were suspended in a 2% gelatin bioink, for bioprinting in a backpressure driven extrusion printhead. The cell bearing bioink was then printed using the same programmed coordinates used for the etching. The bioprinted cells were able to sense and react to the etched features as demonstrated by their elongated orientation along the direction of the etched grooves.

This protocol describes a bioprinting methodology using an automated robotic depositing system that incorporates etched topographical guidance cues with the precision deposition of a cell bearing hydrogel bioink. The printed cells are directly delivered to the etched features and are able to sense and orientate with them.

自动点胶机器人技术在细胞表面和指导生物印刷

This manuscript describes the introduction of cell guidance features followed by the direct delivery of cells to these features in a hydrogel bioink using ...

Synthesis of Hydrogels with Antifouling Properties As Membranes for Water Purification

Hydrogels have been widely utilized to enhance the surface hydrophilicity of membranes for water purification, increasing the antifouling properties and thus achieving stable water permeability through membranes over time. Here, we report a facile method to prepare hydrogels based on zwitterions for membrane applications. Freestanding films can be prepared from sulfobetaine methacrylate (SBMA) with a crosslinker of poly(ethylene glycol) diacrylate (PEGDA) via photopolymerization. The hydrogels can also be prepared by impregnation into hydrophobic porous supports to enhance the mechanical strength. These films can be characterized by attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) to determine the degree of conversion of the (meth)acrylate groups, using goniometers for hydrophilicity and differential scanning calorimetry (DSC) for polymer chain dynamics. We also report protocols to determine the water permeability in dead-end filtration systems and the effect of foulants (bovine serum albumin, BSA) on membrane performance.

This paper reports practical methods to prepare hydrogels in freestanding films and impregnated membranes and to characterize their physical properties, including water transport properties.

水凝胶的合成与防污性能作为膜的净水

Hydrogels have been widely utilized to enhance the surface hydrophilicity of membranes for water purification, increasing the antifouling properties and ...

Surface Engineering of Pancreatic Islets with a Heparinized StarPEG Nanocoating

Cell surface engineering can protect implanted cells from host immune attack. It can also reshape cellular landscape to improve graft function and survival post-transplantation. This protocol aims to achieve surface engineering of pancreatic islets using an ultrathin heparin-incorporated starPEG (Hep-PEG) nanocoating. To generate the Hep-PEG nanocoating for pancreatic islet surface engineering, heparin succinimidyl succinate (Heparin-NHS) was first synthesized by modification of its carboxylate groups using N-(3-dimethylamino propyl)-N'-ethyl carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). The Hep-PEG mixture was then formed by crosslinking of the amino end-functionalized eight-armed starPEG (starPEG-(NH2)8) and Heparin-NHS. For islet surface coating, mouse islets were isolated via collagenase digestion and gradient purification using Histopaque. Isolated islets were then treated with ice cold Hep-PEG solution for 10 min to allow covalent binding between NHS and the amine groups of islet cell membrane. Nanocoating with the Hep-PEG incurs minimal alteration to islet size and volume and heparinization of the islets with Hep-PEG may also reduce instant blood-mediated inflammatory reaction during islet transplantation. This "easy-to-adopt" approach is mild enough for surface engineering of living cells without compromising cell viability. Considering that heparin has shown binding affinity to multiple cytokines, the Hep-PEG nanocoating also provides an open platform that enables incorporation of unlimited functional biological mediators and multi-layered surfaces for living cell surface bioengineering.

This protocol aims to achieve surface engineering of pancreatic islets using a heparin-incorporated starPEG nanocoating via pseudo-bioorthogonal chemistry between the N-hydroxysuccinimide groups of the nanocoating and the amine groups of islet cell membrane.

胰岛血气 StarPEG Nanocoating 的表面工程

Cell surface engineering can protect implanted cells from host immune attack. It can also reshape cellular landscape to improve graft function and ...

Detection of Endotoxin in Nano-formulations Using Limulus Amoebocyte Lysate (LAL) Assays

When present in pharmaceutical products, a Gram-negative bacterial cell wall component endotoxin (often also called lipopolysaccharide) can cause inflammation, fever, hypo- or hypertension, and, in extreme cases, can lead to tissue and organ damage that may become fatal. The amounts of endotoxin in pharmaceutical products, therefore, are strictly regulated. Among the methods available for endotoxin detection and quantification, the Limulus Amoebocyte Lysate (LAL) assay is commonly used worldwide. While any pharmaceutical product can interfere with the LAL assay, nano-formulations represent a particular challenge due to their complexity. The purpose of this paper is to provide a practical guide to researchers inexperienced in estimating endotoxins in engineered nanomaterials and nanoparticle-formulated drugs. Herein, practical recommendations for performing three LAL formats including turbidity, chromogenic and gel-clot assays are discussed. These assays can be used to determine endotoxin contamination in nanotechnology-based drug products, vaccines, and adjuvants.

Detection of Endotoxin in Nano-formulations Using Limulus Amoebocyte Lysate LAL Assays

Detection of endotoxins in engineered nanomaterials represents one of the grand challenges in the field of nanomedicine. Here, we present a case study that describes the framework composed of three different LAL formats to estimate potential endotoxin contamination in nanoparticles.

用利穆鲁布莱巴细胞裂解液 (lal) 法检测纳米配方中的内毒素

When present in pharmaceutical products, a Gram-negative bacterial cell wall component endotoxin (often also called lipopolysaccharide) can cause ...

Wet-spinning-based Molding Process of Gelatin for Tissue Regeneration

This article presents an inexpensive method to fabricate gelatin, as a natural polymer, into monofilament fibers or other appropriate forms. Through the wet spinning method, gelatin fibers are produced by smooth extrusion in a suitable coagulation medium. To increase the functional surface of these gelatin fibers and their ability to mimic the features of tissues, gelatin can be molded into a tube form by referring to this concept. Examined by in vitro and in vivo tests, the gelatin tubes demonstrate a great potential for application in tissue engineering. Acting as a suitable filling gap material, gelatin tubes can be used to substitute the tissue in the damaged area (e.g., in the nervous or cardiovascular system), as well as to promote regeneration by providing a direct replacement of stem cells and neural circuitry. This protocol provides a detailed procedure for creating a biomaterial based on a natural polymer, and its implementation is expected to greatly benefit the development of correlative natural polymers, which help to realize tissue regeneration strategies.

We developed and describe a protocol based on the wet spinning concept, for the construction of gelatin-based biomaterials used for the application of tissue engineering.

明胶湿纺成型工艺促进组织再生

This article presents an inexpensive method to fabricate gelatin, as a natural polymer, into monofilament fibers or other appropriate forms. Through the ...